SUNY at Albany
June 19-23, 2001
High-Resolution Imaging Surface Structures with the Scanning Force Microscope (SFM): A Study on Living Cancer Cells, Fibroblasts and Chondrocytes
SFM studies with different cells, named fibroblast-like synoviocytes (SFC) from rheumatoid joints, chondrocytes, as well as cancer cells from human head-neck squamous cell carcinoma (hypopharynx, UMSCC11 and UMSCC22) and the lung tumor alveolus type (UMSCC7) were done in air and buffer solution. The cells were seeded onto round 4 mm cover slips. Topographic images of untreated cells were studied in serum free media. Considerable tip/surface interactions at individual parts of the cells are observed. The native cells could be influenced by the scanning procedure.
Several observation modes, as non-contact, contact and tapping mode, were tested. In contact mode, briefly described, a tip at the end of a cantilever touches the surface with a constant force and scans line by line the sample. Thereby topographic informations were received by monitoring the change in cantilever deflection. When the tip approach in direct contact to the sample, the living cell induces electronic signals disturbing the scan (Figure 1 right).
Figure. 1: SFM observation of living cells: (left) in non-contact mode the cells are displayed with an unreal softness, (right) in tip/cell surface contact the cell induces electronic signals.
It seems that the living cells are tickled by the scan and avoid further contacts with the SFM-cantilever. By repeating the tip-approach, sometimes the cell dodges or repels the tip. Even after a while, the living cells remember the procedure and avoid another scan. Moving to another part of the sample cannot prevent the repulsion. In so-called "non-contact" mode SFM the cantilever is oscillated at a frequency, which is slightly above the cantilever's resonance frequency typically with amplitudes of a few nanometer. The tip never is brought in physical contact to the sample surface. The two-dimensional scan will be performed only nearby it. The cantilever's resonance frequency is decreased by the van der Waals forces, which extend normally from 1 nm to 10 nm above the adsorbed fluid layer, and by other long-range forces, which extend above the surface. This mode is probably very useful to scan living cells in solution. However, the cells are displayed with an unreal softness (Figure 1 left). Still the general topographic pillow-sized form with intact cell-cell contacts is displayed.
In the present study a ?low force contact mode? was used to scan living cells in liquid medium. A tip with a not too low spring constant (preferential around 0.5 N/m) gave best results to scan the cell. The setpoint was carefully selected to avoid cell damage by the sharp tip. Detail-rich topographic images were observed (Figure 2).
Figure. 2: Magnification of the cell surface (UMSCC22).
The magnification offers highly informative insights into the cell surface. Just as dynamic interactions (e.g. collagen- network assembly) on the surface were monitored. Pores and the protein cover become visible. Some of the pores could be stimulated by the scan. An example of the tip/surface interaction is the pore stimulation of chondrocyte cells. Influenced by the tip the pore changed its energetic potential. Many problems remain to be solved, however, before SFM analysis will routinely deliver reproducible and comprehensive informations for cellular and other biostructures. Therefore, in the present work principally methodological investigations will be intensively introduced.
Gerlinde Bischoff (1), Robert Bischoff (2), Dagmar Riemann (3), Jürgen Langner (3)
Martin Luther University (1) Halle-Wittenberg, Department of Biochemistry/Biotechnology, Kurt-Mothes-Str. 3, 06120 Halle (Saale), Germany
SENSOBI Sensoren GmbH (2), Weinbergweg 22, 06120 Halle, Germnay firstname.lastname@example.org